Electrochemical preparation of 1,1′-dimethylferricinium from a water-soluble 1,1′-dimethylferrocene-2-hydroxypropyl-β-cyclodextrin complex and its applications in enzyme assay

Water-soluble 2-hydroxypropyl-β-cyclodextrin (hp-β-CyD), a cyclic and nonreducing oligosaccharide, was used to enclose the hydrophobic guest molecules ferrocene (FeCp₂) and 1,1′-dimethylferrocene (DMFeCp₂) to form a water-soluble complex. At high concentrations (300 mM), hp-βCyD enclosed up to 100 mM FeCp₂ or DMFeCp₂. The yellow complexes were electrochemically oxidized (platinum vs Ag/AgCl poised at +450 mV) to form the blue dyes ferricinium (FeCp⁺₂) and 1,1′-dimethylferricinium (DMFeCp⁺₂). This is a one-electron transfer process and the ferricinium cations formed exhibited absorption peaks at 620 and 650 nm, respectively. The concentrated DMFeCp⁺₂ was much more stable (4 months) than the FeCp⁺₂ (2 days) and both oxidized dyes were insensitive to a wide pH variation (pH 2-11). The DMFeCp⁺₂ was reduced easily by various reducing agents such as ascorbic acid, uric acid, and sulfite and exhibited an absorption coefficient of 325 cm^-1 M^-1. Application of the novel DMFeCp⁺₂ complex as a colorimetric dye for the enzymatic oxidation of glucose, glutamate, lactate, phenylalanine, xanthine, and hypoxanthine was successfully demonstrated. When applied to real samples the results obtained agreed well with those of standard enzymatic assays. The DMFeCp⁺₂ complex could also be utilized to monitor activity of oxidases under saturating substrate concentrations.